Rapid expression cloning and differential RNA display identifies a gene, prostate tumor inducing gene-1 (PTI-1), that is differentially expressed in prostate cancer versus normal prostate and benign prostatic hypertrophy. PTI-1 encodes a truncated and mutated human elongation factor 1α (EF-1α), and its 5′ untranslated (UTR) region shares significant homology with the 23S ribosomal RNA gene of Mycoplasma hyopneumoniae. PCR with human genomic DNAs, using PTI-1 5′ UTR specific primers, suggests that this sequence is part of the human genome. Furthermore, RT-PCR, with one primer specific to the 5′ UTR region and the other to the EF-1a coding region, amplifies PTI-1 transcripts from total RNA of various human tumor cell lines and blood samples from prostate carcinoma patients. RT-PCR products with the predicted size and sequence of PTI-1 are detected in RNAs from cell lines of human prostate, breast and colon carcinomas. This RT-PCR product is shown by Southern blotting and sequence analyses to contain the junction sequence between the 5′ UTR and the coding region of the PTI-1 gene. Furthermore, RT-PCR analysis indicates that the PTI-1 gene is also expressed in prostate carcinoma patient derived blood samples. On the basis of serial dilution experiments, PTI-1 can detect 1 prostate carcinoma cell in 108 cells not expressing PTI-1. In this context, PTI-1 represents the most sensitive marker currently available for detecting human prostate cancer. This study confirms the authenticity of the PTI-1 gene and documents its potential clinical utility as a sensitive and specific indicator of prostate cancer progression.
Adenocarcinoma of the prostate is presently the most prevalent internal cancer of men in the United States and the second most frequent cause of cancer-related deaths. Current methodologies for the early detection of prostate cancer, including physical examination, monitoring PSA3 levels, tissue biopsy, ultrasound and bone scans, are restricted in both sensitivity and specificity (1-3). In addition, present testing modalities do not permit a distinction between cancers that will remain indolent and those which will prove aggressive and life threatening (1-4). Using DNA transfection approaches with a novel acceptor cell line, CREF-Trans 6 (5), and the molecular approach of differential RNA display (6), a novel putative prostatic carcinoma tumor inducing oncogene, PTI-1, has been identified and cloned from a human prostate carcinoma, LNCaP, cDNA library (7). Using RT-PCR approaches with primers corresponding to the 5′ UTR region of PTI-1, expression is detected in 15 of 16 carcinomas of the prostate, but not in normal prostate or BPH tissue (7). Although further testing with a larger number of patient samples is clearly needed, these provocative results suggest that PTI-1 monitoring might prove beneficial in prostate cancer diagnostics.
The full-length PTI-1 cDNA is 2,123 bp and it encodes a truncated and mutated human EF-1α (7) (FIG. 1). The structure of the PTI-1 cDNA is unique in that its 5′ UTR shares significant homology (approximately 85%) with the prokaryotic 23S ribosomal RNA gene from Mycoplasma hyopneumoniae. This high degree of sequence homology between the 5′ UTR of the PTI-1 gene and prokaryotic 23S ribosomal RNA gene raises concerns that contamination by bacteria in the LNCaP cell culture used to prepare the cDNA library and subsequent cloning artifacts may be responsible for the identification of the PTI-1 gene. Confirming the authenticity of the PTI-1 gene is mandatory before further studies can be conducted to elucidate any potential role of PTI-1 in human prostate cancer development and evolution.
In the present study, the question of the validity of the PTI-1 gene was addressed by analyzing its presence in the human genome, transcripts in tumor cell lines and presence in blood samples from patients with prostate cancer. The results of these investigations demonstrate definitively that the identification of the PTI-1 gene is unlikely due to bacterial contamination and/or technical artifacts. Moreover, PTI-1 gene expression may provide an extremely sensitive marker for prostate carcinoma progression as reflected by the presence of prostate carcinoma cells in a patients' bloodstream.